Genetic recombination in Streptomyces griseus.

Low-frequency (10(-6)) genetic recombination was observed in a cephamycin-producing strain of Streptomyces griseus. The recombinants were predominantly heteroclones. Heteroclone analysis was performed involving four heteroclones of one cross. In 100 mutants correlation was found between the type of auxotrophy and the level of antibiotic activity. A cross of this strain with a streptomycin-producing strain of S. griesus is described.     

Genetic recombination in Nocardia mediterranei.

The regulation of macromolecular biosynthesis was studied in a temperature-sensitive mutant of Escherichia coli previously identified as containing a single mutation causing a thermolabile sn-glycerol-3-phosphate acyltransferase, the first enzyme of the pathway for phospholipid biosynthesis. When this mutant was shifted to a nonpermissive temperature, phospholipid synthesis, as well as ribonucleic acid, deoxyribonucleic acid, and protein synthesis, decreased in a coordinate manner, suggesting the existence of a common regulatory mechanism. During the same time that the rate of macromolecular synthesis was decreasing at the nonpermissive temperature, the intracellular concentration of adenosine 5'-triphosphate dropped dramatically and the concentration of adenosine monophosphate increased. The concentration of adenosine 5'-diphosphate dropped, but not as markedly. The decrease in macromolecular synthesis and the changes in the adenine nucleotide concentrations can now be attributed to a thermolabile adenylate kinase. The inactivation of adenylate kinase prevented the cell from converting adenosine 5'-monophosphate to adenosine 5'-diphosphate and consequently from making adenosine 5'-triphosphate. This in turn caused a decrease in the rate of macromolecular synthesis and cell growth. Adenylate kinase, therefore, is a key enzyme in controlling the rate of cell growth. The nature of the possible relationship between adenylate kinase and glycerol-3-phosphate acyltransferase is discussed.     

Genetic analysis of recombination in prokaryotes.

Bacteria provide a simple system for the genetic analysis of homologous recombination. More than twenty genes have been identified in Escherichia coli. The enzymatic activities associated with the products of many of these genes have been revealed by studies with model DNA substrates. It is now possible to pair homologous molecules in vitro and process these through defined intermediates into mature recombinants of the types predicted by genetic crosses.     

Genetic transformation and transfection ofBacillus subtilis spheroplasts

Genetic transformation and transfection of lysozyme-treatedBacillus subtilis spheroplasts 168M ind occurs only if they are stabilized with 0.5m phosphate buffer and not if they are stabilized with 0.5m sucrose. Spheroplasts prepared from maximally competent cells give maximum transformation and transfection results. The results indicate that the DNA receptors must also be intact in the spheroplasts.     

Genetic recombination of human immunodeficiency virus.

We investigated genetic recombination of the human immunodeficiency virus (HIV) in a tissue culture system. A clonal cell line expressing a single integrated HIV provirus with a termination codon affecting pol gene expression was transfected with different defective mutants derived from an infectious molecular clone of HIV. Replication-competent viral particles were recovered, passaged, and plaque purified. Restriction analyses of the proviral DNA corresponding to several of these viruses indicated that their emergence was the result of genetic recombination.     

Genetic recombination in plants

Meiotic recombination generates novel allelic arrays on chromosomes. Recent experiments have revealed an extraordinarily nonrandom distribution of recombination breakpoints along the lengths of plant chromosomes; for example, recombination breakpoints often resolve within genic sequences, and thereby generate novel alleles. The mechanism by which recombination breakpoints are determined is an area of active investigation. In addition, recent developments are providing recombination-based technologies for creating targeted alterations in the architecture of plant genomes.     

Genetic recombination in Actinoplanes brasiliensis by protoplast fusion.

Protoplast formation, fusion, and cell regeneration have been achieved with mutant strains of Actinoplanes brasiliensis. Three-, four-, and five-factor crosses have shown genetic recombination among the markers, and a five-factor cross is analyzed and discussed. Possibilities of using protoplast fusion for gene mapping and strain improvement are suggested.     

Genetic recombination in Micromonospora rosaria by protoplast fusion.

Auxotrophic strains of Micromonospora rosaria were isolated by N-methyl-N'-nitro-N'-nitrosoguanidine mutagenesis and used in intraspecific recombination by protoplast fusion. High-frequency fusion of protoplasts of M. rosaria strains was induced by polyethylene glycol (molecular weight, 1,000) (PEG 1,000). The optimum concentration of PEG 1,000 for fusion of M. rosaria was 50% (wt/vol). PEG 4,000 was slightly better than PEG 1,000 at concentrations lower than 50% (wt/vol). The recombinant frequency did not increase after treatment with PEG 1,000 (50% [wt/vol]) for longer than 20 min. Under these conditions, fusion with many auxotrophic strains of M. rosaria resulted in a high frequency of formation of true recombinants (sometimes more than 10%). Additionally, when ros (rosamicin nonproducing) strains were crossed by protoplast fusion; about 5% of the resultant prototrophic recombinants were shown to have the ros+ (rosamicin producing) characteristic restored. Rosamicin production by M. rosaria colonies was clearly distinguished by the broth overlay method. The results of fusion experiments between ros and ros+ strains indicated that either the chromosomal mutation or pleiotrophic effect of some auxotrophic markers is involved.